title: noninfected_m9
description: Protocols: RNA was extracted (see nucleic acid extraction protocol) from freshly extracted small intestinal epithelium (see below) from mice infected with Nippostrongylus brasiliensis and not (see treatment protocol), a library was created (see nucleic acid library construction protocol) and sequenced paired end 150 bp (see nucleic acid sequencing protocol) Epithelium from small intestine was isolated using slightly modified version of a published protocol (Sato 2013). In summary, the small intestine of mouse was washed in ice-cold PBS and opened longitudinally. A microscopy cover slip was used to gently scrape of excess mucus and villus before the intestine was cut into 2-4 mm pieces. The fragments were resuspended in 30 mL ice-cold PBS and pipetted up and down with a 10 mL pipette. Supernatant was discarded and this step was repeated 5-10 times until the supernatant was clear. The fragments were incubated in 2 mM EDTA in PBS at 4 C for 30 min with gentle rocking. The supernatant was removed, 20 mL of ice-cold PBS was added and the fragments were pipetted up and down. This washing step was repeated until the crypt fraction appeared as seen in a microscope. The 1-3 consecutive fractions were collected into a FCS coated 50 mL tube. References: Sato, Toshiro and Hans Clevers (2013). “Primary Mouse Small Intestinal Epithelial Cell Cultures”. In:Epithelial Cell Culture Protocols: Second Edition. Ed. by Scott H. Randell and M. Leslie Fulcher. Methods in Molecular Biology. Totowa, NJ: Humana Press, pp. 319-328.isbn: 978-1-62703-125-7.doi:10.1007/978-1-62703-125-7_19.527url:https://doi.org/10.1007/978-1-62703-125-7_19. Mice were infected with Nippostrongylus brasiliensis RNA was extracted with a column based RNA extraction kit such as Qiagen spin miniprep. Library preparation was done with NEB Next(R) Ultra(TM) RNA Library Prep Kit
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2021-05-01T00:23:35Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2020-06-03T20:20:23Z
attributes.tag: External Id
attributes.value: SAMEA6872461
attributes.tag: INSDC center name
attributes.value: Dept. of Clinical and Molecular Medicine Norwegian University of Science and Technology (NTNU)
attributes.tag: INSDC first public
attributes.value: 2021-05-01T00:23:35Z
attributes.tag: INSDC last update
attributes.value: 2020-06-03T20:20:23Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-9183:noninfected_m9
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: common name
attributes.value: house mouse
attributes.tag: developmental stage
attributes.value: adult
attributes.tag: genotype
attributes.value: wild type genotype
attributes.tag: individual
attributes.value: H
attributes.tag: organism part
attributes.value: epithelium of small intestine
attributes.tag: sample name
attributes.value: E-MTAB-9183:noninfected_m9
attributes.tag: scientific_name
attributes.value: Mus musculus
attributes.tag: strain
attributes.value: C57BL/6